Silver nanoparticles (AgNP), the most popular nano-compounds, possess unique chemical, physical and biological properties. Albizia adianthifolia (AA) – rich in saponins – is a plant of the Fabaceae family, found abundantly on the East coast of Africa. This plant is well known for its medicinal properties, and although the exact phytochemistry of AA is unknown, recent research suggests that AA can be used for the treatment of certain pathologies. The biological properties of a novel silver nanoparticle (AAAgNP) synthesised from an aqueous leaf extract of AA, were investigated on A549 lung carcinoma cells. Cell viability was determined by the 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay. Cellular oxidative status (lipid peroxidation and glutathione (GSH) levels) were determined by the TBARS and GSH-Glo™ Glutatione assays respectively. ATP concentration was measured using the CellTitre-Glo™ assay. Caspase-3/-7, -8 and -9 activities were determined by Caspase-Glo® assays. Flow cytometry was used to measure apoptosis, mitochondrial (mt) membrane depolarisation, expression of CD95 receptors and intracellular smac/DIABLO levels. DNA fragmentation was assessed with the comet assay. The expression of p53, bax, PARP-1 and smac/DIABLO was evaluated by western blotting. Quantitative polymerase chain reaction was used to determine mRNA levels of bax and p53. AAAgNP caused a dose-dependent decrease in cell viability with a significant increase in lipid peroxidation (5-fold vs. control; p=0.0098) and decreased intracellular GSH (p=0.1184). A significant 2.5-fold decrease in cellular ATP was observed upon AAAgNP exposure (p=0.0040) with a highly significant elevation in mt membrane depolarisation (3.3-fold vs. control; p<0.0001). Apoptosis was also significantly higher (1.5-fold) in AAAgNP treated cells (p<0.0001) with a significant decline in expression of CD95 receptors (p=0.0416). AAAgNP caused a significant 2.5-fold reduction in caspase-8 activity (p=0.0024) with contrasting increases in caspase-3/-7 (1.7-fold vs. control; p=0.0180) and -9 activity (1.4-fold vs. control; p=0.0117). Western blots showed increased expression of smac/DIABLO (4.1-fold) in treated cells (p=0.0033). Furthermore, AAAgNP significantly increased the expression of p53, bax cleaved PARP-1 (1.2-fold; p=0.0498, 1.6-fold; p=0.0083 and 1.1-fold; p=0.0359 respectively). The expression of mRNA for both p53 and bax was also elevated post AAAgNP treatment, with 6-fold (p=0.0036) and 5-fold (p=0.0080) changes respectively compared to untreated cells. Data suggests that AAAgNP induces cell death in the A549 lung cells via the mt-mediated intrinsic apoptotic program. Further investigations are required to assess the potential use of AAAgNP in cancer treatment. / Thesis (M.Med.)-University of KwaZulu-Natal, Durban, 2012.

Nanotoxicology is a nascent field of study concerned with the potential for nanotechnology to adversely impact human health or result in ecological damage. Nanomaterials can display unique physicochemical properties not present in the parent bulk material and it is these properties that may be a potential source of toxicity. There are a growing number of examples of nanomaterials functioning differently in biosystems compared to the parent bulk material. With the rapid growth of nanotechnology and increasing exposure of people to novel nanomaterials there is an urgent need to evaluate the toxicity of nanomaterials. In this study the toxicities of silver and zinc oxide nanoparticles were assessed. The effects of size and surface coating on the cytotoxicity and immunogenicity of silver nanoparticles were investigated, with cytotoxicity found to be inversely proportional to nanoparticle size. The subcutaneous penetration of zinc oxide nanoparticles was assessed to determine whether this material can be safely used as a UV filter in sunscreens and cosmetics. No dermal penetration was detected using a porcine in vitro model. Zinc oxide nanoparticles were also used as a model material to investigate nano-specific toxicity by comparing cytotoxicity and changes to gene expression with bulk scale zinc oxide. In both cases cytotoxicity and changes to gene expression were greater for zinc oxide nanoparticles. Methods and techniques to test the toxicity of nanomaterials in vitro and the implication for in vivo toxicity are only beginning to be elucidated. The methods and techniques used in this study, particularly nanomaterial stabilization in biofluids and toxicity testing using blood cell cultures, may assist the establishment of standard in vitro testing protocols for nanomaterials.